Biotechnology

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Biotechnology Questions

Bring on the tough stuff

1. Scientists use restriction enzymes isolated from bacteria for DNA cloning. How do bacteria prevent restriction enzymes from dicing up their own DNA?

2. You have been working on cloning the gene you study into a cloning vector and think you finally have it. You ramp up production of the recombinant DNA in E. coli and isolate the DNA. What mechanisms can you use to ensure that the plasmid you have isolated from your bacteria is what you were expecting?

3. Being a typical teenager, you spend most warm summer days outside. Mosquitos? Who needs insect repellent? You're a tough cookie! A few days after enjoying a spectacular Fourth of July fireworks display you start to feel sick. You head to your doctor, who, after a quick exam, wants to have you tested for West Nile virus. West Nile is an RNA virus. What test or technique can be used to confirm whether or not you indeed have West Nile?

4. Your chemistry buddies have just synthesized a new small molecule that may have therapeutic potential. You have no idea what this molecule does, so you'd like to start out by examining its effect on gene expression. Which technique are you most likely to use and why?

5. What advantages do stem cells have over any other type of cell for scientific research?

6. Dolly the sheep was the first mammal to be cloned using the nucleus from a differentiated cell. The nucleus used was from a six year old sheep. Unfortunately, she started to develop health issues normally seen in much older sheep and was eventually euthanized at a young age. In fact, many cloned animals seem prone to health issues. Provide an explanation for why cloned animals see the onset of health issues at a young age?

7. You are working as a forensic scientist and have a DNA sample from the crime scene as well as samples from three suspects. Describe how you would determine the guilty suspect using this evidence.

8. What is gene therapy? What are the potential problems with this type of treatment?

9. What are induced pluripotent cells? What advantage do they have over embryonic stem cells?

10. Your teacher gives you a sample of all the inside components of a human cell (cell extract) and asks you to identify whether or not the sample contains the protein, dicer. What technique will you use and why?

Possible Answers

1. Scientists use restriction enzymes isolated from bacteria for DNA cloning. How do bacteria prevent restriction enzymes from dicing up their own DNA?

Restriction enzymes exist in bacterial cells as a defense mechanism to cut up any foreign DNA. They protect their own genomes by adding methyl groups to cytosine and adenine nucleotides.

2. You have been working on cloning the gene you study into a cloning vector and think you finally have it. You ramp up production of the recombinant DNA in E. coli and isolate the DNA. What mechanisms can you use to ensure that the plasmid you have isolated from your bacteria is what you were expecting?

First, most cloning vectors contain some sort of antibiotic resistance gene. This ensures that when you grow your bacteria in the antibiotic, only the bacteria that have taken up your recombinant DNA with the antibiotic resistance gene remain. (The others should be killed off). Secondly, you can digest the DNA with the same restriction enzymes you used to create your recombinant DNA and use gel electrophoresis to separate the fragments and ensure they are the sizes you expected and that you don't see anything unexpected. Finally, you can sequence your recombinant DNA to make sure that it is the exact sequence you expect.

3. Being a typical teenager, you spend most warm summer days outside. Mosquitos? Who needs insect repellent? You're a tough cookie! A few days after enjoying a spectacular Fourth of July fireworks display you start to feel sick. You head to your doctor, who, after a quick exam, wants to have you tested for West Nile virus. West Nile is an RNA virus. What test or technique can be used to confirm whether or not you indeed have West Nile?

Since West Nile virus is caused by an RNA virus, reverse transcriptase polymerase chain reaction (RT-PCR) can be used. RT-PCR uses the enzyme, reverse transcriptase to convert RNA to DNA. Therefore, presence of a product would indicate that the mRNA was present.

4. Your chemistry buddies have just synthesized a new small molecule that may have therapeutic potential. You have no idea what this molecule does, so you'd like to start out by examining its effect on gene expression. Which technique are you most likely to use and why?

You would most likely use DNA microarray analysis. You could use Southern blotting, but you would have to go gene by gene, and with the advent of microarrays, this is really impractical. Microarray analysis will give you genes or family of genes that may be affected by this small molecule.

5. What advantages do stem cells have over any other type of cell for scientific research?

Stem cells are advantageous because they are not differentiated meaning they have the potential to become any type of cell. Other types of cells are already differentiated meaning they have already become specialized.

6. Dolly the sheep was the first mammal to be cloned using the nucleus from a differentiated cell. The nucleus used was from a six year old sheep. Unfortunately, she started to develop health issues normally seen in much older sheep and was eventually euthanized at a young age. In fact, many cloned animals seem prone to health issues. Provide an explanation for why cloned animals see the onset of health issues at a young age?

To clone a mammal, scientists take a differentiated cell and dedifferentiate it in the lab. This dedifferentiated nucleus is used to clone the animal. It could be possible that the nucleus has not been fully dedifferentiated. The fact that the nucleus used to clone Dolly was from a six year old sheep could have contributed to the fact that aged prematurely.

7. You are working as a forensic scientist and have a DNA sample from the crime scene as well as samples from three suspects. Describe how you would determine the guilty suspect using this evidence.

The simplest way to analyze this data would be to analyze the short tandem repeats for each sample. You would use primers for a particular STR and perform PCR on each sample. Following PCR, you would use gel electrophoresis and compare the samples of your suspects to the DNA left at the crime scene. Suspect samples with products of different size to the crime scene DNA can be eliminated, while those with the same size are most likely guilty.

8. What is gene therapy? What are the potential problems with this type of treatment?

Gene therapy is a technique that delivers a properly functioning gene to a cell whose gene is mutated or absent. In theory, some problems associated with gene therapy include the following: the effect is not long lasting, it introduces something foreign into the cell which can trigger an immune response, the viral vectors used to deliver the gene can be toxic, and finally, many diseases are the result of several genes, and gene therapy can only target single genes.

9. What are induced pluripotent cells? What advantage do they have over embryonic stem cells?

Induced pluripotent cells are already differentiated but have been coaxed in the lab to become pluripotent. This means, like embryonic stem cells, they can become any type of cell. The advantage of these cells is that the way they are obtained does not raise any ethical red flags. Unfortunately, scientists have uncovered some differences between embryonic stem cells and induced pluripotent cells.

10. Your teacher gives you a sample of all the inside components of a human cell (cell extract) and asks you to identify whether or not the sample contains the protein, dicer. What technique will you use and why?

Break out the compass and start walking. Just kidding! You would perform a Western blot. A Western blot looks for the presence of a particular protein.